Zacharoff, Lori2016-12-192016-12-192016-09https://hdl.handle.net/11299/183383University of Minnesota Ph.D. dissertation. September 2016. Major: Biochemistry, Molecular Bio, and Biophysics. Advisor: Daniel Bond. 1 computer file (PDF); xii, 140 pages.Iron is the fourth most abundant element in the Earth’s crust. Part of the biogeochemical cycling of iron occurs in stratified ecological zones in the anoxic layer in a process aided by dissimilatory metal reducing bacteria. Geobacter sulfurreducens is a model organism for this respiratory process. This organism harnesses respiratory energy from the metabolism of substrates with a low amount of available free energy. Since metal reduction occurs extracellularly, metabolic electrons have to be transferred distances that are measured in micrometers. The final step of metal respiration requires G. sulfurreducens to make direct contact with extracellular metals and then transfer electrons to extracellular metals through insulating membrane barriers. The metals respired add another level of complexity to respiration. For example, insoluble iron oxides change electrical potential and degree of solubility while being reduced. Currently, the molecular mechanism of the metal reducing processes has yet to be elucidated but multiheme cytochromes are known to be involved. This dissertation focuses on the inner membrane cytochrome CbcL. The goal was to find a protein that is capable of coupling the movement of electrons to the generation of a proton motive force in the inner membrane. Physiological and electrochemical approaches described in this dissertation support the conclusion that CbcL is an inner membrane protein important for extracellular metal respiration. CbcL functions in a way that is specific to the redox potential of the terminal electron acceptor. This provides a mechanism for precisely coupling energy harvesting to the amount of free energy provided by the extracellular terminal electron acceptor. Further, this dissertation contains evidence that the final step of electron transfer to metals involves the protein PgcA. Using biochemical, electrochemical and phenotypic analyses, PgcA adheres to insoluble metals, has apparent electron shuttling properties and is required by G. sulfurreducens only for reduction of insoluble metals and not soluble ferric citrate or poised electrodes.enThesis or Dissertation